Systematic functional analysis of the Com pilus in Streptococcus sanguinis : a minimalistic type 4 filament dedicated to DNA uptake in monoderm bacteria

Abstract

ABSTRACT Type 4 filaments (T4F) are a superfamily of versatile nanomachines, ubiquitous in prokaryotes, which use conserved multi-protein machineries to assemble and operate filamentous polymers of type 4 pilins. In the best-studied T4F, these machineries are complex, which has posed challenges to understanding the mechanisms of filament assembly and their multiple functions. Here, we report the systematic functional analysis of the Com pilus, a widespread T4F mediating DNA uptake during natural transformation in monoderm bacteria. Using Streptococcus sanguinis , we show that Com pili are bona fide type 4 pili (T4P), representing a new pilus sub-type. We show that with only eight components necessary for pilus assembly and functioning—all “core” proteins universally conserved in this superfamily—the Com pilus epitomizes a minimalistic T4F. We further demonstrate that core T4F components are sufficient for filament assembly. Paradoxically, akin to the more elaborate T4F, the Com pilus contains four minor pilins forming a complex, probably tip-located. Our results have global implications for T4F and make the Com pilus a model for elucidating fundamental processes underpinning filament assembly. IMPORTANCE Type 4 filaments (T4F) are nanomachines ubiquitous in prokaryotes, centered on filamentous polymers of type 4 pilins. T4F are exceptionally versatile and widespread virulence factors in bacterial pathogens. The mechanisms of filament assembly and the many functions they facilitate remain poorly understood because of the complexity of T4F machineries. This hinders the development of anti-T4F drugs. The significance of our research lies in characterizing the simplest known T4F—the Com pilus that mediates DNA uptake in competent monoderm bacteria—and showing that four protein components universally conserved in T4F are sufficient for filament assembly. The Com pilus becomes a model for elucidating the mechanisms of T4F assembly.